In 1980, Jaeken et al. (Pediatric Research 14:179) described a new neurological disorder in twin girls. Since then over 200 similar patients have been described, mostly in Northern Europe, but the syndrome is found in various countries. Patients present in the neonatal period with hypotonia, failure to thrive, and dysmorphic features including esotropia, inverted nipples, and subcutaneous fat over the suprapubic region. Severe mental and psychomotor retardation with hepatomegaly, cerebellar hypoplasia and brain stem atrophy are seen. Many have vomiting, diarrhea, coagulopathy, stroke-like episodes, pericardial effusions, seizures, and retinitis pigmentisa (McDowell and Gahl (1997) Proc. Soc. Exp. Biol. Med. 215: 145-157; Krasnewich and Gahl (1997) Advances in Pediatrics 44: 109-140; Jaeken et al. (1993) Glycobiology 3: 423-428; Kristiansson et al. (1998) J. Pediatr. Gastroenterol. Nutr. 27: 23-29). A comprehensive monograph describing clinical features, stages, progression and biochemical analysis appeared in 1991 (Jaeken et al. (1991) Acta Paediatr. Scand. Suppl. 375: 1-71).
The syndrome reported by Jaeken et al. was the first of a new group of multisystemic disorders caused by faulty glycosylation that is now emerging. These Carbohydrate-Deficient Glycoprotein Syndromes (CDGS) are usually neuropathies, but non-neurological forms of CDGS have appeared recently (Niehues et al. (1998) J. Clin. Invest. 101:1414-1420; de Koning et al. (1998) Biochem Biophy Res Commun 245:38-42; Jaeken et al. (1998) Amer. J. Hum. Genet. 62: 1535-1539). Some cases present with hypoglycemia, severe protein-losing enteropathies, persistent vomiting along with coagulopathy. All CDGS patients are believed to have primary defects that directly affect the N-glycosylation pathway.
The most commonly used method for diagnosing CDGS is a transferrin isoelectric focusing (IEF) test, which was originally used to screen for alcoholism. Serendipitously, the transferrin IEF test was found to be useful for diagnosing some of the CDGS disorders. The basis for this test is that transferrin usually has two complex-type N-linked oligosaccharide chains, each with two sialic acids, for a total of four negative charges. A few oligosaccharide chains have three sialic acids, yielding transferrin molecules with five and six negative charges. Any genetic or physiological condition that reduces the number of sugar chains on the proteins, or changes the structure of the sugar chains so they carry fewer sialic acids, will likely change the isoelectric point and the IEF pattern. Most known cases of CDGS result in loss of an entire sugar chain. A few patients make incomplete chains. Both give rise to partially carbohydrate deficient transferrin (CDT) with an altered IEF pattern. The transferrin IEF test forms the basis for biochemically dividing the CDG syndromes into four types (Henry et al. (1997) J. Lab. & Clin. Med. 129: 412-421; Stibler et al. (1993) Neuropediatrics 24: 51-52; Stibler et al. (1995) Neuropediatrics 26: 235-237; Jaeken et al. (1993) J. Inherit. Metab. Dis. 16: 1041). Analysis of the isoforms of other serum glycoproteins can also be used to diagnose CDGS. More than 40 different serum glycoproteins have been shown to have altered isoforms or decreased activities (Harrison et al. (1992) Clin. Chem. 38: 1390-1392; Henry et al. (1997) J. Lab. & Clin. Med. 129: 412-421; Stibler et al. (1998) Scand. J. Clin. Lab. Invest. 58: 55-61). These include antithrombin III (AT-III), factor XI and protein C.
Evidence suggests that the already identified CDG syndromes are just the tip of the iceberg of diseases and other conditions that are due to misglycosylations. For example, the preliminary findings by Murch et al. that symptoms and pathologies of inflammatory bowel disease are improved by providing N-acetylglucosamine to the patients are provocative (Murch et al. (1998) British Society of Gastroenterology (Abstract)). The mechanism, of course, is unknown, and neither “glycoscience” nor medicine have a data base that provides satisfactory explanations. Additional areas for further investigation for both clinicians and glycobiologists have been suggested (Gahl W A. (1997) J. Lab. Clin. Med. 129: 394-395; Kornfeld S (1998) J. Clin. Invest. 101: 1293-1295).
Approximately fifty percent of dysmorphic and dysfunctional children that appear at the pediatric clinic remain undiagnosed throughout childhood and adult life. Many are broadly diagnosed with “seizure disorders,” “cerebral palsy,” or as “mentally retarded.” Some have early deaths. Aberrant glycosylation is seldom suspected as a cause of these conditions or other human diseases, even though approximately one percent of human genes encode proteins that produce and/or recognize oligosaccharides.
The discovery and treatment of additional diseases that are associated with deficiencies in glycosylation has been hampered by a lack of efficient and effective diagnostic tests. The IEF assays of transferrin and other serum glycoproteins suffer from significant drawbacks as diagnostic tools for CDGS and other glycosylation disorders. For example, a specific IEF pattern does not define a specific mutation, as any one of several different defects can give rise to the same IEF pattern. In the case of transferrin, three different biochemical defects all yield the same “Type I” transferrin IEF pattern. A second disadvantage is that transferrin contains only a small number of oligosaccharide structures, so the IEF test can identify only a small fraction of glycosylation-deficient states. Moreover, temporary physiological conditions such as uncontrolled fructosemia, galactosemia, and recent heavy alcohol consumption can also disturb glycosylation and produce abnormal transferrin IEF patterns (Stibler et al. (1997) Acta Paediatrica 86: 1377-1378; Jaeken et al. (1996) Pediatric Research 40:764-766). Yet another major shortcoming is that fewer than five diagnostic laboratories in the US analyze serum transferrin IEF mobility.
The lack of adequate methods for diagnosing glycosylation disorders is particularly tragic in that once recognized, life-threatening defects in glycosylation can be treated and sometimes cured. In at least one case, for example, a CDGS patient was successfully treated with a dietary therapy of the sugar mannose. Therefore, a need exists for simple, efficient, and accurate methods for diagnosing glycosylation disorders. The present invention fulfills this and other needs.